Project Summary Our knowledge of mechanisms regulating vascular remodeling in atherosclerotic and non- atherosclerotic vasculopathies is still inadequate. This represents an important rate-limiting step to improve or design new therapeutic interventions that may prove beneficial during atherogenesis or in advanced atherosclerotic lesions. In the cardiovascular system, globins including hemoglobin and myoglobin are thought exclusively to transport and store molecular oxygen and regulate nitric oxide bioavailability. However, recent evidence suggests that this model is not complete and mammalian globins such as cytoglobin (CYGB) might have important functions in vascular remodeling that are independent of oxygen storage and transport. We discovered that CYGB is abundantly expressed in medial smooth muscle cells in the vessel wall of rodent and human vessels. We have obtained strong preliminary results indicating that CYGB inhibits apoptosis during vascular injury in vivo and in vitro. The goal of this proposal is to test the general hypothesis that smooth muscle CYGB combines novel anti-apoptotic and nitric oxide consuming functions that have favorable effects in inhibiting atherosclerotic plaque pathogenesis. In Aim 1, we will test the hypothesis that smooth muscle specific conditional knockdown of CYGB will increase smooth muscle cell apoptosis, with a detrimental loss of smooth muscle cells, and a decrease in indices of plaque stability. We will use novel genetically encoded cell lineage tracing approaches in a mouse model of atherosclerosis combined with targeted conditional deletion of smooth muscle CYGB. In Aim 2, we will establish the contribution of nitric oxide signaling to the function of CYGB in the vasculature and in advanced atherosclerotic plaques through functional and metabolomics characterization. In Aim 3, we will determine the mechanism of action of CYGB in apoptosis. To this end, we will establish the role of CYGB in regulating the downstream effector of apoptosis caspase-3 through redox- and gene-regulated inhibition of its activation. Overall, this work will be guided through studies of human atherosclerotic plaque samples derived from asymptomatic and symptomatic patients. The results derived from the proposed studies will challenge the current dogma related to mammalian globin functions as simply oxygen transport and nitric oxide handling systems. Ultimately, we hope to delineate novel pro-survival pathways in the vasculature that could be manipulated to mitigate inappropriate vascular remodeling through combined regulation of apoptosis and nitric oxide bio-availability.